Regulation of endosome fusion

Homotypic fusion between early endosomes can be reconstituted in vitro. By using wortmannin and LY294002, inhibitors of phosphatidylinositol (Pl) 3-kinase, a requirement for this activity has been established in order for fusion to proceed efficiently. It has been shown that Pl 3-kinase activity is required downstream of rab5 activation, although a large excess of activated rab5 can overcome wortmannin inhibition. A series of experiments have also been performed which indicate a role for early endosomal autoantigen 1 (EEA1) in determining fusion efficiency. EEA1 dissociates from membranes following wortmannin treatment. It is proposed that the requirement of endosome fusion for Pl 3-kinase activity is to promote the association of EEA1 with endosomes.

Inhibition of endosome fusion by wortmannin persists in the presence of activated Rab5

Rab5-dependent endosome fusion is sensitive to the phosphoinositide 3-kinase inhibitor, wortmannin. It has been proposed that phosphoinositide 3-kinase activity may be required for activation of rab5 by influencing its nucleotide cycle such as to promote its active GTP state. In this report we demonstrate that endosome fusion remains sensitive to wortmannin despite preloading of endosomes with stimulatory levels of a GTPase-defective mutant rab5(Q79L) or of a xanthosine triphosphate-binding mutant, rab5(D136N), in the presence of the nonhydrolysable analogue XTPgammaS. These results suggest that activation of rab5 cannot be the principal function of the wortmannin-sensitive factor on the endosome fusion pathway. This result is extrapolated to all GTPases by demonstrating that endosome fusion remains wortmannin sensitive despite prior incubation with the nonhydrolysable nucleotide analogue GTPgammaS. Consistent with these results, direct measurement of clathrin-coated vesicle-stimulated nucleotide dissociation from exogenous rab5 was insensitive to the presence of wortmannin. A large excess of rab5(Q79L), beyond levels required for maximal stimulation of the fusion assay, afforded protection against wortmannin inhibition, and partial protection was also observed with an excess of wild-type rab5 independent of GTPgammaS.

Do Children still need to Escape Childhood? A Reassessment of John Holt and his Vision for Children's Rights

The work of children’s liberationists have been long been critiqued for pushing the parameters of rights discourse too far; specifically, by suggesting that there are no significant differences between children and adults, including their ability for self-determination. John Holt’s 1974 text Escape from Childhood is one such work which was deemed highly controversial for its time. This article uses Holt’s Escape from Childhood as an overarching framework against which to examine the current state of play on children’s rights as explicated through the UN Convention on the Rights of the Child. It suggests that whilst Holt has often been critiqued for being too radical, in the context of current children’s rights discourse Holt’s visioning is not as radical as it might first appear.

Escape and connection : a phenomenological investigation into the meaning of an after-school program for adolescent boys

The purpos e of this phenomenological research is to explore the meaning of a YMCA-sponsored after-school recreation program in the lives of four adolescent boys. Listening to youth voice is impor t ant to the ability of othe r s to design, implement and evaluate high-quality programs tha t facilitate learning opportunities tha t a r e meaningful to participants. Within the context of interviews, task-based activities we r e used to ga the r data. Guided by Creswell's analytic spiral (1998), data wa s analyzed according to van Manen's (1990) thematic analysis and Caeilli's (2000) creative narrative analysis. It wa s found tha t this after-school progr am provided the s e adolescents with the opportunity to escape from the i r monotonous after-school activities and the instability of the i r home and school environments. Also, they we r e connected wi th positive peers, caring adults and the wide r community, opportunities tha t we r e limited in othe r aspects of the i r lives. Methodological issues a r e also discussed.

Modelado y control de recirculación de gases de escape en motores a inyección de combustible

Tesis (Maestro en Ciencias de la Ingeniería Eléctrica con Especialidad en Control) UANL, 2000.

Implication de l'endosome de recyclage dans la migration cellulaire in vivo

Au cours de l’ovogenèse chez la mouche du vinaigre: Drosophila melanogaster, un groupe de cellules folliculaires appelées cellules de bord, migrent à travers les cellules nourricières pour atteindre l’ovocyte. Cet événement, nécessitant la transition épithélio- mésenchymateuse (TEM), la réorientation, puis l’arrêt, ressemble à la formation de métastases. L’endocytose est un régulateur clé de plusieurs événements polarisés, y compris la migration cellulaire. En effet, différentes protéines impliquées dans la migration, comme les intégrines et les E-cadhérines (cadhérines épithéliales), sont régulées par transport à travers les endosomes. De même, l’endocytose restreint au front de migration l’activité des récepteurs tyrosine kinases (RTKs) qui guident les cellules de bord dans leur mouvement. Cependant les mécanismes moléculaires de cette restriction spatiale de l’activité des RTKs demeurent largement inconnus. Nous avons testé l’implication du trafic vésiculaire à travers la machinerie d’endocytose, dans la migration dirigée des cellules de bord, car ce système est facilement accessible pour l’expression de protéines et l’analyse de mutants. Nous avons commencé par confirmer une observation précédente du rôle de l’endosome précoce dans la migration des cellules de bord. Ensuite, nous avons identifié l’endosome de recyclage (ER) comme un régulateur clé de cette migration. En effet, nous avons démontré que l’expression dans les cellules de bord d’une forme dominante négative de Rab11, la petite GTPase régulant le transport vésiculaire à travers l’ER, bloque la migration ou entraîne de sévères défauts de migration dans environ 80% des chambres d’œufs examinées. De plus, nous observons par immunofluorescence une relocalisation de l’activité des RTKs alors que d’autres protéines de migration ne sont pas affectées par Rab11 dominant négatif. Ce résultat a été par la suite confirmé par une interaction génétique entre Rab11 et les RTKs. D’autre part, nous avons montré que le complexe exocyste, un effecteur de Rab11, est impliqué dans la migration des cellules de bord. Nous avons trouvé par microscopie confocale en tissu fixé et par microscopie en temps réel que Sec15, un composant de ce complexe, est polarisé, de façon Rab11- dépendante, dans des vésicules qui s’accumulent au front de migration tout au long du mouvement des cellules de bord. De plus, la perte de l’activité de Sec15 perturbe à son tour la migration. Ainsi, toutes ces données démontrent le rôle fondamental d’un cycle d’endo- exocytose dans le maintien des RTKs actifs au niveau du front de migration des cellules de bord le long de leur mouvement.

Micelles polyioniques ternaires pour la libération intracellulaire d’oligonucleotides

Les oligonucléotides (ONs) antisens présentent un fort potentiel en tant qu’agents thérapeutiques. Toutefois, leurs propriétés physicochimiques limitent leur utilisation en thérapie génique. Pour pallier aux divers obstacles, des systèmes de vectorisation, tels que les micelles polyioniques (PICMs), ont été développés. Grâce à leur structure unique, les micelles protégent l’ON contre une dégradation prématurée et le couplage d’un ligand à leur surface augmente leur spécificité et leur internalisation. Dans d’autres systèmes, un polymère adjuvant aux propriétés pH-sensibles peut être ajouté pour faciliter la sortie de l’endosome et augmenter l’efficacité de l’ON. L’objectif général de ce mémoire était de mettre au point des PICMs ternaires ciblées pour l’administration d’ONs. Ces micelles assureraient à la fois l’internalisation cellulaire de leur cargaison en interagissant avec des récepteurs cellulaires et sa fuite de l’endosome grâce à un mécanisme de déstabilisation de la membrane endosomale. Pour cela, des PICMs composées d’un copolymère cationique de type poly(éthylène glycol)-bloc-poly(méthacrylate d’(alkylamino)éthyle) et d’un copolymère d’acide méthacrylique ont été préparées. Les propriétés physicochimiques de ces vecteurs ont démontré qu’ils permettaient une condensation efficace de l’acide nucléique et ce, indépendamment de la nature du polymère cationique et de l’acide nucléique. Finalement, une approche de couplage par pont disulfure a été développée afin de greffer au copolymère un fragment d’anticorps dirigé contre les récepteurs de la transferrine. En conclusion, ces travaux démontrent la versatilité et le potentiel des PICMs ternaires en tant que vecteurs d’acide nucléique, et proposent une méthodologie de couplage d’un ligand afin de formuler des PICMs ciblées.

The Greek debt trap: an escape plan. Bruegel Policy Contribution 2012/19, 7 November 2012

Without corrective measures, Greek public debt will exceed 190 percent of GDP, instead of peaking at the anyway too-high target ratio of 167 percent of GDP of the March 2012 financial assistance programme. The rise is largely due to a negative feedback loop between high public debt and the collapse in GDP, and endangers Greek membership of the euro area. But a Greek exit would have devastating impacts both inside and outside Greece. A small reduction in the interest rate on bilateral loans, the exchange of European Central Bank holdings, buy-back of privately-held debt, and frontloading of some privatisation receipts are unlikely to be sufficient. A credible resolution should involve the reduction of the official lending rate to zero until 2020, an extension of the maturity of all official lending, and indexing the notional amount of all official loans to Greek GDP. Thereby, the debt ratio would fall below 100 percent of GDP by 2020, and if the economy deteriorates further, there will not be a need for new arrangements. But if growth is better than expected, official creditors will also benefit. In exchange for such help, the fiscal sovereignty of Greece should be curtailed further. An extended privatisation plan and future budget surpluses may be used to pay back the debt relief. The Greek fiscal tragedy highlights the need for a formal debt restructuring mechanism

Very short delays prior to escape from potential predators may function efficiently as adaptive risk-assessment periods

Periods between predator detection and an escape response (escape delays) by prey upon attack by a predator often arise because animals trade-off the benefits such a delay gives for assessing risk accurately with the costs of not escaping as quickly as possible. We tested whether freezing behaviour (complete immobility in a previously foraging bird) observed in chaffinches before escaping from an approaching potential threat functions as a period of risk-assessment, and whether information on predator identity is gained even when time available is very short. We flew either a model of a sparrowhawk (predator) or a woodpigeon (no threat) at single chaffinches. Escape delays were significantly shorter with the hawk, except when a model first appeared close to the chaffinch. Chaffinches were significantly more vigilant when they resumed feeding after exposure to the sparrowhawk compared to the woodpigeon showing that they were able to distinguish between threats, and this applied even when time available for assessment was short (an average of 0.29 s). Our results show freezing in chaffinches functions as an effective economic risk assessment period, and that threat information is gained even when very short periods of time are available during an attack.

Internationalization motives: sell more, buy better, upgrade and escape

Purpose – The purpose of this the paper is to review the motives for internationalization to clarify previous arguments and provide a theory-driven classification. Design/methodology/approach – The authors build on behavioral economics and propose a classification of internationalization motives as the result of the interaction among two dimensions, an economics-driven exploitation of existing resources or exploration of new resources, and a psychology-driven search for better host country conditions or avoidance of poor home country conditions. Findings – These two dimensions result in four internationalization motives: sell more, in which the company exploits existing resources at home and obtains better host country conditions; buy better, in which the company exploits existing resources abroad and avoids poor home country conditions; upgrade, in which the company explores for new resources, and it obtains better host country conditions; and escape, in which the company explores for new resources and avoids poor home country conditions. Originality/value – This theory-driven classification provides predictive power for future analyses of internationalization motives.

Triple- and quadruple-escape peaks in HPGe detectors: Experimental observation and Monte Carlo simulation

The triple- and quadruple-escape peaks of 6.128 MeV photons from the (19)F(p,alpha gamma)(16)O nuclear reaction were observed in an HPGe detector. The experimental peak areas, measured in spectra projected with a restriction function that allows quantitative comparison of data from different multiplicities, are in reasonably good agreement with those predicted by Monte Carlo simulations done with the general-purpose radiation-transport code PENELOPE. The behaviour of the escape intensities was simulated for some gamma-ray energies and detector dimensions; the results obtained can be extended to other energies using an empirical function and statistical properties related to the phenomenon. (C) 2010 Elsevier B.V. All rights reserved.